The development of the nervous system requires the proper differentiation, migration and morphogenesis of neurons. The morphological differentiation of individual neurons and assembly of the trillions of neuronal connections that compose the human nervous system occurs through guided extension of axons and dendrites. Molecular guidance cues in the environment of developing neurons guide neuronal growth cones at the tips of extending axons and dendrites. mTOR-mediated local synthesis of new proteins within growth cones as emerged as an important mechanism that controls axon guidance. Mutations in genes involved local protein synthesis are responsible for several human autism spectrum disorders, including Fragile X syndrome and Tuberous Sclerosis Complex (TSC). While modulation of mTOR-dependent protein synthesis is known to be required downstream of both attractive and repulsive axon guidance in several animal model systems, it is unknown if similar mechanisms function in developing human neurons. This proposal will first test whether human Retinal Ganglion Cells (RGCs) derived from human induced pluripotent stem cells (hiPSCs) use mTOR-mediated protein synthesis to respond to positive and negative axon guidance cues. In Aim 2 we will test the role of the TSC1/TSC2 complex, which is a key upstream negative regulator of mTOR. For this we will generate new lines of hiPSCs by reprograming fibroblast cells from patients with TSC. While this proposal will focus on RGCs, a wide variety of other cell and neuronal types can be studied using these new hiPSC lines. Therefore, these important new cell lines will be a valuable resource for many investigators and will be made available for distribution through WiCell. In Aim 3 we will test whether RGCs derived from TSC hiPSCs exhibit abnormal response to axon guidance cues tested in Aim 1.